Upon activation by mitogen activated protein kinases (MAPKs), c-Jun undergoes phosphorylation, which affects its DNA binding activity and stability. The underlying mechanism and the key enzymes responsible for this phenomenon remain largely unknown. In particular, the phenomenon of c-Jun degradation in the inflammatory response has not yet been reported. To verify this, we investigated LPS-stimulated murine macrophages pre-treated with sodium orthovanadate (SO) to uncover the regulatory mechanism of the MAPKs that regulate c-Jun degradation in the inflammatory response. SO suppressed the production of prostaglandin E2 (PGE2) and the expression of COX-2 in LPS-stimulated RAW264.7 cells. SO also decreased total c-Jun levels without altering the abundance of its mRNA, although phospho-levels of p38, ERK, and JNK were strongly enhanced. By using selective MAPK inhibitors and knockdown and overexpression strategies, p38 was revealed to be a major MAPK that regulates c-Jun degradation. Further analysis indicated that the phosphorylation of p38 is a determinant for c-Jun degradation that is sufficient to induce ubiquitination-dependent c-Jun degradation, recovered by MG132 treatment. Therefore, our results suggest that hyperphosphorylation of p38 by SO contributes to c-Jun degradation linked to the suppression of PGE2 secretion in inflammatory responses; finding drugs to increase p38 activity could be a novel strategy for the development of anti-inflammatory drugs.